We propose to study the formation of gap junctions/low resistance junctions between Novikoff hepatoma cells, correlating quantitative data obtained with ultrastructural (thin section and freeze-fracture) and physiological (ionic and dye transfer) methods. Employing computer-assisted analysis, we will examine cells dissociated with EDTA and reaggregated for varying lengths of time, with and without a previous recovery period. These studies will provide data on rates of formation and stages in junctional development. To further elucidate the mechanisms of formation, we will analyze: junctional permeability in single pairs of cells manipulated into contact, protease effects on junctional development between reaggregating cells, cholesterol distributions in formation plaques, and the results of quick-freeze EM methods on the structure of mature and developing junctions with known permeabilities. We will also attempt to modify junction formation by varying temperature, altering membrane lipid composition, treating with agents that affect negative surface charges and applying cyclic nucleotide derivatives. We expect these studies to increase our understanding of the structure and dynamics of the plasma membrane, to give additional insight into the regulation of cell junctions, to provide new tools for testing the biological significance of junctional communication, especially in the control of cell proliferation, and to offer further basis for evaluating the possible importance of altered junction formation in cancer or other pathological states.